The acetylation of lysine residues by histone acetyltransferases regulates protein function. Cells contain multiple, distinct families of histone acetyltransferases, including Tip60. Tip60 is implicated in the pathogenesis of several human disease processes, including cancer progression, neurodegenerative diseases and viral replication. The participation of Tip60 in these diverse disease processes reflects Tip60's ability to regulate transcription, viral replication and the cells response to DNA damage. Tip60 regulates the cells response to genotoxic stress through its ability to acetylate and activate the ATM protein kinase. Loss of ATM function is associated with increased incidence of cancer, sensitivity to ionizing radiation and cerebellar neurodegeneration. Tip60 is therefore a key regulator of the cells ability to repair and survive DNA damage. Radiation therapy is a major treatment modality for human cancer. However, many tumor types are relatively resistant to the cytotoxic effects of radiation therapy. Accordingly, selective pharmacological inhibition of Tip60 provides a potentially powerful therapeutic target for enhancing the efficacy of radiotherapy and chemotherapy in the treatment of human tumors. Currently, no selective inhibitors of histone acetyltransferases in general, and particularly of Tip60, have been identified. Thus, in this proposal we propose to develop a high throughput assay to identify novel, specific inhibitors of the Tip60 histone acetyltransferases. 2 specific aims will be undertaken. In specific aim 1, a novel high throughput ELISA assay will be developed which utilizes the ability of recombinant Tip60 to acetylate immobilized peptides. In specific aim 2, the assay will be formatted to a 384-well format, and preliminary screening of active library compounds will be carried out. Tip60 represents a target for which an inadequate array of selective and potent small molecule inhibitors exist; screening and identifying such inhibitors would have a significant impact in studying the role of Tip60 and related HATs in the progression of human disease. This proposal will develop specific inhibitors of the Tip60 HAT which can be employed to disrupt cellular protein acetylation levels with the aim of improving treatment of human tumors. These inhibitors can be used in animal and clinical studies to test if Tip60 [unreadable] In activation sensitizes human tumors to radiation therapy and chemotherapy. In addition, Tip60 inhibitors can be employed to probe the basic signaling mechanisms by which Tip60 exerts its biological effects, and to identify new Tip60 substrates. Finally, Tip60 inhibitors can be used in complex organisms, such as mice, to examine the contribution of Tip60 to disease processes such as cancer progression, neurodegenerative diseases and genotoxic stress. [unreadable] [unreadable] [unreadable]